open access

Vol 46, No 2 (2008)
Original paper
Submitted: 2011-12-19
Published online: 2008-06-04
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Triple immunofluorescence labeling of atherosclerotic plaque components in apoE/LDLR -/- mice.

Mariusz Gajda, Jacek Jawień, Lukasz Mateuszuk, Grzegorz J Lis, Andrzej Radziszewski, Stefan Chłopicki, Jan A Litwin
DOI: 10.2478/v10042-008-0021-8
·
Folia Histochem Cytobiol 2008;46(2):143-146.

open access

Vol 46, No 2 (2008)
ORIGINAL PAPERS
Submitted: 2011-12-19
Published online: 2008-06-04

Abstract

This paper presents a simple and reliable method of triple immunofluorescence staining that allows simultaneous detection of various cell types present in atherosclerotic plaque of apolipoprotein E and LDL receptor-double knockout (apoE/LDLR -/-) mice. We used combined direct and indirect procedures applying commercially available primary antibodies raised in different species to detect smooth muscle cells (Cy3-conjugated mouse anti-smooth muscle actin, SMA), macrophages (rat anti-CD68) and T lymphocytes (rabbit anti-CD3). Fixation of the material in acetone and modified incubation protocol employing nonfat dry milk in preincubation and incubation media significantly increased the intensity of labeling and effectively quenched the background. Our method offers an efficient way to detect qualitative as well as quantitative changes of macrophages, T lymphocytes and smooth muscle cells in atherosclerotic plaque of apoE/LDLR -/- mice during atherosclerosis development or in response to pharmacological treatment.

Abstract

This paper presents a simple and reliable method of triple immunofluorescence staining that allows simultaneous detection of various cell types present in atherosclerotic plaque of apolipoprotein E and LDL receptor-double knockout (apoE/LDLR -/-) mice. We used combined direct and indirect procedures applying commercially available primary antibodies raised in different species to detect smooth muscle cells (Cy3-conjugated mouse anti-smooth muscle actin, SMA), macrophages (rat anti-CD68) and T lymphocytes (rabbit anti-CD3). Fixation of the material in acetone and modified incubation protocol employing nonfat dry milk in preincubation and incubation media significantly increased the intensity of labeling and effectively quenched the background. Our method offers an efficient way to detect qualitative as well as quantitative changes of macrophages, T lymphocytes and smooth muscle cells in atherosclerotic plaque of apoE/LDLR -/- mice during atherosclerosis development or in response to pharmacological treatment.
Get Citation
About this article
Title

Triple immunofluorescence labeling of atherosclerotic plaque components in apoE/LDLR -/- mice.

Journal

Folia Histochemica et Cytobiologica

Issue

Vol 46, No 2 (2008)

Article type

Original paper

Pages

143-146

Published online

2008-06-04

DOI

10.2478/v10042-008-0021-8

Bibliographic record

Folia Histochem Cytobiol 2008;46(2):143-146.

Authors

Mariusz Gajda
Jacek Jawień
Lukasz Mateuszuk
Grzegorz J Lis
Andrzej Radziszewski
Stefan Chłopicki
Jan A Litwin

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